Nonspecific Defenses Against Microbial Infection

I. For the most part, pathogenic organisms are held at bay not by the immune system but by several nonspecific barriers and mechanisms. These nonspecific means of limiting the invasion of the body by microorganisms act to keep bacteria and fungi from colonizing those sites within the body in which they would flourish.

II. Virtually all surfaces of your body that are directly exposed to the environment are covered by either skin or mucous membranes.

A. The integument or skin covers the external part of our body. It presents a physical barrier to the movement of pathogens into the nutrient rich environment of the underlying tissues. The uppermost layer of the skin is referred to as the epidermis. The epithelial cells of this tissue are loaded with the protein known as keratin. Keratin serves to toughen and waterproof these cells. The cells of the epidermis are held together by protein interactions between the cells. The tightness of the junctions between cells is a physical barrier to the invasion of the underlying tissues.

1. Sebaceous glands found in the epidermis secrete an oily substance known as sebum which lubricates the skin and exerts a mild antimicrobial effect. Sweat glands not only secrete sweat but also an enzyme known as lysozyme which disrupts the cell wall of gram positive organisms. The drying of sweat on the surface of the skin changes the osmotic condition of the skin radically and thus limits the growth of those organisms which are unable to withstand such changes.
2. The combination of the secretions found on the skin surface and the action of commensalate organisms living on the skin cause the pH of this surface to be below 5. This also exerts another nonspecific barrier to the growth of many organisms.
3. Because of the reduced load of microorganisms on the surface of the skin, minor abrasions and cuts will often heal without complicating infections. It should be noted that stress, hormonal shifts (as in puberty) can result in alteration in the secretions of the sebaceous glands. This in turn can result in the opportunistic infection of the skin and glands of the skin.

B. Underlying the epidermis is the dermis. In most sites the dermis consists of a relatively thick layer of dense fibrous connective tissue. This dense tissue consists of (among other things) fibers of the protein collagen and a substance which consists of a mixture of amino acids and polysaccharides known as hyaluronic acid. The dermis with its dense packed nature, provides a further physical barrier to the spread of microorganisms. Unlike the epidermis, the dermis is vascular so certain inflammatory processes occur here that do not occur in the epidermis. Furthermore, macrophages (known as histiocytes) will phagocytize invading bacteria and mast cells which help amplify the reaction to an invading bacteria allow for a second line of defense in the dermis. These will be discussed shortly.

C. Lining all internal surfaces that are exposed to the environment (GI tract, urinary tract, reproductive tract and respiratory tract) are the mucous membranes. The cells of these membranes are arranged into two distinct layers. The upper layer isa   tightly joined layer of epithelial cells similar to epidermis but unlike the epidermis in that they are not waterproofed or toughened with keratin. The underlying layer of tissue is a loose connective tissue. The lack of keratin and the loose nature of the connective tissue leave the mucus membrane susceptible to dehydration. Glands found in these membranes secrete mucus which covers the cells to help minimize water loss. The warm, moist environment of the mucous membranes are ideal for colonization by pathogens. In most sites microbial antagonism plays an important role in limiting infection with pathogenic organisms.

1. Mucus can trap bacteria. In the respiratory tract bacteria are constantly being moved out of the lower parts of the trachea by the action of ciliary escalator. This mechanism moves the bacteria trapped in the mucus up to the back of the throat where it can be swallowed. The acids of the stomach act to destroy the bacteria.
2. Other substances are found in the secretions of the mucous membranes which exert a negative impact on the ability of bacteria to colonize these membranes. Acids secreted in the stomach, lysozyme and antibodies (IgA) secreted along the digestive tract are but a few of the barriers to bacterial colonization.

III. Before we can discuss the second or third line of defense you will need to be introduced to the white blood cells (WBC’s or leukocytes). WBC's can be divided into two general groups, the granulocytes (neutrophils, eosinophils and basophils) and the agranulocytes (monocytes and lymphocytes).  When stained using a procedure known as Wright’s staining, granulocytes have numerous granules in the cytoplasm.  Agranulocytes usually lack such granules. These cells are made and released from the bone marrow into the blood stream.  They circulate throughout the body and leave the bloodstream to enter tissues where they are responsible for surveillance of these tissues.  Once released from the bone marrow a leukocytes will not last remain in the bloodstream long.  Consequently, there is a constant need to replenish the circulating leukocytes. Often the presence of an invading pathogen triggers an increased rate of leukocyte release from the bone marrow.  This leads to elevation in the number of circulating WBC's.     

A) Neutrophils are the most abundant leukocyte, constituting approximately 55%-65% of the circulating leukocytes.  Neutrophils are active early in most bacteria infections.  Using phagocytosis, neutrophils will internalize and destroy bacterial cells.  Neutrophils are also capable of releasing substances that will kill bacteria and will attract more WBC’s.

B) Eosinophils are far less common, constituting 1-3% of the total number of leukocytes.  These cells are capable of phagocytosis but do not seem to carry this out very often. They seem to play an important role in phagocytizing antigen-antibody complexes.  The granules of the eosinophil contain chemicals that will kill parasitic worms. These granules are released in response to infection by parasitic worms and help the body rid itself of these worms. The role of the eosinophil in the inflammatory response is less well understood.  Under certain conditions these cells play an important proinflammatory role while under other conditions they seem to release substances that reign in the inflammatory response.  

C) The basophil is the rarest of the WBC’s constituting 0.5% -1% of the total number of white cells.  The granules of the basophil contain histamine and heparin.  When released these chemicals increase edema and vasodilation. 

D) The monocyte constitutes approximately 3-7% of the WBC’s. The monocyte is recruited to the site of infection late in the inflammatory response.  When they leave circulation and enter the tissue they are referred to by a number of names including macrophage, histiocyte, Kupffer cells (in the liver) and dendritic cells.  These cells are highly active  phagocytes. Once the materials have been phagocytized they are fragmented and the pieces returned to the surface of the macrophage or dentritic cells. These fragments are presented to the lymphocytes (antigen presentation). For most lymphocytes this is a required step in their activation. Dendritic cells (DC's) migrate from the tissue where they phagocytized materials into the lymph node that is part of the lmphatics that drain the tissue.  DC's are extremely important in engaging T lymphocytes (T cells) within the lymph node  to become active in the fight against infection.  

E) The lymphocytes make up 25-33% of the WBC’s. The are generally divided into T lymphocytes (T cells) and B lymphocytes (B cells).  Roles played by the lymphocytes are quite diverse and will be covered in a later lecture in more detail.  Briefly, they are responsible for production of antibodies when they mature into plasma cells and for identification and destruction of virus infected cells.  

IV. If a pathogen breaches the barriers of the epithelial layer of either the skin or the mucous membranes and colonizes the underlying connective tissues, a second line of nonspecific defense is brought to bear. For the most part this involves the reticuloendothelial system (RES). Remember that the majority of tissue are supported by a mesh of connective tissues referred to as reticular connective tissue. This tissue not only supports the specialized cells of various tissues and organs but plays an important role in surveillance. Within the RES are many cells that are capable of phagocytosis. These cells known as phagocytes recognize and ingest pathogen in a process known as phagocytosis. Several types of phagocytic cells are involved; both neutrophils, those cells derived from monocytes (macrophages and dendritic cells) and to a lesser extend  eosinophils (eosinophils are capable of phagocytosis but does not appear to play a significant role in clearing bacteria in this manner). Both neutrophils and monocytes are transported by the blood stream and leave circulation to enter tissue throughout the body. This movement out of the blood stream by the white blood cell is referred to as diapedesis. Along with the RES, several other systems must be mentioned with regard to body defense. The lymphatic system also plays an important role in body defense. It consists of a system of thin walled vessels that drain the extracellular fluids from tissues. The fluid, known as lymph, passes through lymph nodes in which many cells of the immune system are harbored. Dendritic cells migrate from the tissues, where they have ingested foreign proteins, to the lymph nodes, where they will present the foreign proteins to lymphocytes.  

A. The phagocyte is attracted to the pathogen due to chemicals released from the organism itself or proteins released due to the damage caused by the infection. This is referred to as chemotaxis. The pathogen is attached to the membrane of the phagocyte which is referred to as adherence. At that point the pathogen is ingested by the extension of phogocytic cell's cell membrane out and around the pathogen. The pathogen is contained within a membrane bound vacuole known as a phagosome.   A lysosome fuses with the phagosome (forming a phagolysosome) and the pathogen is broken down by the action of the lysosomal enzymes.

B. In the case of the neutrophil, this is the end of the story. But dendritic cells and  macrophages will digest the pathogen and then bring little pieces of the proteins that are specific to that organism to their surface. This is referred to as antigen presentation and is crucial in the activation of some aspects of our immune response.

V.  Inflammation occurs when the trauma to or infection of a tissue occurs.    The hallmarks of inflammation are swelling (edema), localized heat (calor), redness (rubor) and pain (dolor).  In the case of inflammation associated with trauma to a tissue, the trauma damages cells;  these damaged cells initiate the inflamatory response.   When a pathogen colonizes a tissue it will often cause the tissue damage.  The damaged cells and /or bacteria themselves  release substances  that cause the clinically apparent inflammatory response.  Inflammation associated with infection with bacteria serves to limit the spread of and eventually destroy the pathogenic agent.  One of the important chemicals activated early in the inflammatory response is bradykinin.  This protein is found in body fluids in an inactive state.  Damage to the cell's of a tissue leads to release of substances from the damaged cells which  activate bradykinin in the extracellular fluids of the damaged tissue.  

A.   Important in the early progression of the inflammatory response are the  mast cells.  These cells are  found in the submucosa or dermis.  Histamine and seretonin (among other things) are released from the mast cells due in part to the pressence of activated bradykinin.   Initially, bradykinin, histamine and serotonin contribute to a  transient vasoconstriction.  This is followed by dilation of the arterioles  in the region that has been affected.  This is referred to as vasodilation.    The histamine causes a formation of gaps between the endothelial cells of the capillaries and venules.  This loss of integrity results in edema due to the flooding of the tissue with fluids from the blood stream.

B.  Bradykinin and other substnaces produced during inflammation will alter the surface of endothelial cells causing WBC's to attach to the interior lining of the blood vessel.  This causes an accumulation of WBC's within the blood vessel in the inflamed tissue, a condition known as margination.   

C.  During inflamation, several types of cells are capable of converting phospholipids into prostaglandin and leukotrienes.  Prostaglandin and leukotrienes are responsible for initiating and maintaining a wide range of inflammatory changes within an affect tissue. Many anti-inflammatory drugs work by blocking the production of one or both of these substances.

D.   White blood cells which were involved in margination, migrate from the vessels into the injured or infected tissues.   This is referred to as diapedesis.  Usually, neutrophils are the first cells to leave circulation and begin phagocytosis.  If they encounter bacteria, the neutrophil will release substances that recruit more WBC’s to the site.  If the inflammatory reaction is sustained monocytes leave circulation (becoming macrophages).

E.  The pus seen in some inflammatory reactions is due to the accumulation of dead white blood cells (mainly neutrophils) at the site of infection. Pus is rarely seen in inflammatory responses triggered by trauma alone.

F. Often infection results in fever. Many bacteria and some viruses will release chemicals or cause the macrophages which have ingested them to release chemicals that will result in the hypothalamus' allowing the body's temperature to rise. The lipopolysaccharide found as part of the outer membrane of gram negative organisms will strongly trigger this response. The chemicals released from the macrophage which trigger the fever are generally referred to as endogenous pyrogens while chemicals released from bacteria or viruses that directly cause fever are referred to as exogenous pyrogens. 

VI. Along with the chemicals released by cells during the inflammatory response are several other chemicals secreted by cells which have been infected with viruses or by the lymphocytes that have been specifically activated.

A. Interferon alpha and beta are proteins produced by cells when they are infected with a virus. The release of interferon into the environment is a signal to surrounding cells that viruses are present. This results in subtle changes in the neighboring cells that render them less susceptible to infection by the virus.

B. Interleukins are released by macrophages (interleukin 1) or helper T cells (interleukin 2) and have specific effects on other lymphocytes or in the case of interleukin 1, play a role in stimulating the hypothalamus to increase body temperature.

 

VII.  One other nonspecific defense is the complement system.  The proteins  (roughly 30 distinct proteins) circulate in the blood and are found in the interstitial fluids.  These proteins are  activated by the binding of antibodies (we'll talk about them shortly) to the surface of a bacteria, by the interaction of the certain proteins of the complement system with the proteins on the surface of a gram negative bacteria, or by the binding of  proteins released by the liver to the surface of a bacterial cell.   Activation of the system results in the rupturing of the bacteria cells, vasodilation and increased chemotaxis and margination.

A.  The classical pathway involves the recognition of a bacteria by an antibody.  The antibody binds to the bacteria specifically.  This allows   complement protein 1 (C1) to bind to the antibody-antigen complex.  C1 binds C4 and C2.  The combination of C1, C2 and C4  activates C3 by clipping a portion of the protein off.   The small cleaved portion (C3a) attracts more WBC’s to the area.   C3b (the larger protion of C3) then activates C5 by cleaving it into two fragments.  C5a increase the intensity of the inflammatory response while C5b begins the process of assembling the membrane attack complex ( which consist of C6-C9;  it is referred to as MAC).  The MAC punches holes in hte membrane of bacteria.
B.  The alternative pathway  relies upon the binding of complement proteins  to the surface of certain bacteria directly.  Directly binding of complement usually begins with the binding of C3 to the surface of a bacterial cell.  Activation of the complement cascade occurs in a similar manner as that described above. The outcome of this activation is virtually the same as with the classical pathway.
C.  The liver release certain proteins referred to as the acute phase proteins  in response to infection.  These proteins affect the body in several ways.  They are capable of  binding to the surface of certain bacteria and  activating the complement cascade to destroy the bacteria.

Want to know more?  Here are some good links!!!

Medical Microbiology Textbook  Immunology Overview  http://gsbs.utmb.edu/microbook/ch001a.htm

Univesity of Connecticut  Body Defenses http://www.sp.uconn.edu/~terry/229sp98/immunesys.html